SARS-CoV2 in public spaces in West London, UK during COVID-19 pandemic

Background Spread of SARS-CoV2 by aerosol is considered an important mode of transmission over distances >2 m, particularly indoors. Objectives We determined whether SARS-CoV2 could be detected in the air of enclosed/semi-enclosed public spaces. Methods and analysis Between March 2021 and December 2021 during the easing of COVID-19 pandemic restrictions after a period of lockdown, we used total suspended and size-segregated particulate matter (PM) samplers for the detection of SARS-CoV2 in hospitals wards and waiting areas, on public transport, in a university campus and in a primary school in West London. Results We collected 207 samples, of which 20 (9.7%) were positive for SARS-CoV2 using quantitative PCR. Positive samples were collected from hospital patient waiting areas, from hospital wards treating patients with COVID-19 using stationary samplers and from train carriages in London underground using personal samplers. Mean virus concentrations varied between 429 500 copies/m3 in the hospital emergency waiting area and the more frequent 164 000 copies/m3 found in other areas. There were more frequent positive samples from PM samplers in the PM2.5 fractions compared with PM10 and PM1. Culture on Vero cells of all collected samples gave negative results. Conclusion During a period of partial opening during the COVID-19 pandemic in London, we detected SARS-CoV2 RNA in the air of hospital waiting areas and wards and of London Underground train carriage. More research is needed to determine the transmission potential of SARS-CoV2 detected in the air.


Liquid-based Total Suspended Particulate (TSP) Samplers
Unsegregated total suspended particulates (TSP) were collected using the commercially-available liquid-based samplers including the Biospot-VIVAS (Aerosol Devices Inc., Ft. Collins, CO), the Sartorius MD8 (Sartorius AG, Germany) and the Coriolis μ air sampler (Bertin Technologies). The Biospot VIVAS has been shown to be efficacious in collecting viral particles [11] and airborne particles using a water vapour condensation method, at a rate of 8 litres per minute (L min -1 ). Air initially passes through a cool temperature conditioner, followed by passage through the initiator at 30°C which coalesces particles as small as 8 nm into larger droplets greater than 2 µm in diameter. The enlarged particles are then collected through a set of nozzles onto 1.5 mL of liquid collection media. The collection media used, composed of 1.5mL 1x phosphate buffered saline (PBS), 0.5% (w/v) bovine albumin fraction V and sucrose added to a final concentration of 0.2M, has been used to successfully culture SARS-CoV-2 [1]. Liquid samples were stored at 4°C and transported on ice for viral RNA extraction and SARS-CoV-2 PCR testing.
The Sartorius MD8 (Sartorius AG, Germany) sampled air at a rate of 30 L min -1 for 30 minutes, impacting particles onto a sterile gelatin filter (80 mm diameter, 3 μm pore size, type 80-ACD, Sartorius AG). After sampling, the gelatin filter was transferred aseptically into a Petri dish and transported on ice and stored in a refrigerator at 4°C for later processing.
We also used the Coriolis μ air sampler (Bertin Technologies) which collects air at 100 L min -1 . Samples were collected every 30 minutes into a conical vial containing 15 mL phosphate-buffered saline. Samples were stored at 4°C with viral RNA extraction fluid and qRT-PCR performed on neat samples. All the filters used in the above-mentioned size segregated particle samplers were allowed to equilibrate in a weighing room with controlled temperature (21°C) and relative humidity (30-40%) for a minimum of 24 hr prior to weighing before and after sampling. After each sampling session, filters were removed using forceps and inserted into sterile Petri dishes and sealed using parafilm. These were transported on ice to a level 2 laboratory for particle extraction and SARS-CoV-2 PCR testing.

Portable Samplers
We also sampled using the portable SKC Button sampler (SKC LTD, Dorset, UK). Air was extracted at a rate of 4 L min -1 using the SKC Airchek touch pump, which was attached to the outlet of the button or a filter cassette. loaded with a 37mm PTFE filter. The sampler was attached to an individual's clothing, on their chest, ~2 cm under their clavicle, whilst wearing an FFP3 mask. Devices were calibrated using an adaptor by attaching the Button sampler to the inlet of the button, and the inlet of the calibration adaptor to a HVAC system prior to use. Air samples were collected directly onto a dissolvable 25 mm gelatin filter (1.0μm pore size, SKC LTD, Dorset) when sampling with the button, or unto the 37mm PTFE filter (0.3μm pore size, SKC LTD, Dorset) preloaded to a filter cassette, using an appropriate adaptor as per manufacturer's instructions. The button sampler collects 'inhalable' particles with a 1μm-100µm aerodynamic diameter as defined by the British Standards Institution (BS EN 481:1993). The PTFE loaded cassette collects total suspended particulate with an aerodynamic diameter above 0.3µm.

PUF and TF filters
Filters were removed from air samplers using forceps and placed in a Petri dish at the sampling site. PBS was added to the filters in a class II biological safety cabinet (10 ml to polyurethane foam (PUF), 5 ml to PFTE) and sealed with parafilm. This was then placed on a shaker for 30 minutes at 70 RPM to gently transfer particles from the filter into the solution. After 30 minutes, the solution was transferred into a 10 ml Falcon tube in an L2 HEPA-filtered hood using a pipette. 1 ml of the solution was immediately sent for culture on Vero-E6 cells in a 1.5 ml Eppendorf, with the remaining solution stored at -80ºC for RNA extraction and RT-qPCR.

Gelatin filters
80 mm gelatin filters were placed into a Petri dish with 10 ml of PBS added whereas 37 mm gelatin filters from portable samplers were placed in 50 ml falcon tubes with 5 ml of PBS added. They were then placed in a shaker at 70 RPM and warmed to 28ºC for 30 minutes to dissolve the filter. The remaining solution was transferred into a 10 ml Falcon tube in a L2 HEPA-filtered hood using a pipette. 1 ml of the solution was immediately sent for culture on Vero-E6 cells in a 1.5 ml Eppendorf tube, with the remaining solution stored at -80 ºC for RNA extraction and RT-qPCR.

Biospot VIVAS and Coriolis
Air particles from the Biospot VIVAS were directly collected in liquid medium in a small petri dish and conical flask respectively. After sampling, an equal volume of the solution (between 500-1000 μl) was transferred into three separate 1.5ml Eppendorf tubes. One Eppendorf tube containing solution was immediately sent for culture on Vero-E6 cells, with the remaining stored at -80ºC for RNA extraction and RT-qPCR.